The rational development of new antineoplastic agents directed against tubulin, a protein critical for cell division, requires greater understanding of the interactions between the polypeptide subunits of tubulin and its two tightly bound guanine nucleotides. Interactions of ribose-modified GDP and GTP analogs with tubulin were examined in both microtubule-associated protein (MAP) dependent and independent systems. These studies continue to indicate that GDP and GTP interact with tubulin by different mechanisms. The analogs 2', 3'-dideoxyguanosine 5'-di- and triphosphate (ddGDP and ddGTP) in particular had unique properties: ddGTP supported polymerization without MAPs under conditions in which GTP and all other analogs were inert; and ddGDP, but no other diphosphate, supported vigorous microtubule formation in a reaction requiring MAPs. A clinically useful assay for taxol was developed, and preliminary pharmacokinetics were performed. NSC-181928, a methotrexate analog with antimitotic activity, was demonstrated to be an effective antitubulin agent. Studies on the ADP-ribosylation of tubulin by cholera toxin were initiated. Efforts to separate the alpha and beta subunits of tubulin continued.